Abstract
The location of the dissimilatory nitrite reductase and orientation of its reducing site of the Grampositive denitrifier, Bacillus firmus NIAS 237 were examined. Approximately 90% of the total dissimilatory nitrite reductase activity with ascorbate-reduced phenazine methosulfate (PMS) as the electron donor was on the protoplast membrane. Nitrite induced with intact Bacillus cells an alkalinization in the external medium, followed by acidification. The electron transfer inhibitor, 2-heptyl-4-hydroxyquinoline-N-oxide, which blocked nitrite reduction with endogenous substrates, inhibited the acidification, but not the alkalinization. Alkalinization was not affected with ascorbate-reduced PMS as the artificial electron donor. This indicated that the alkalinization is not associated with proton consumption outside the cytoplasmic membrane by the extracellular nitrite reduction. The dissimilatory nitrite reductase of B. firmus NIAS 237 was located on the cytoplasmic membrane, and its reducing site is suggested to be on the inner side of this membrane.
Similar content being viewed by others
Abbreviations
- CCCP:
-
carbonylcyanide m-chlorophenylhydrazone
- HOQNO:
-
2-heptyl-4-hydroxyquinoline-N-oxide
- PMS:
-
phenazine methosulfate
- H+/NO sup-inf2 ratio:
-
number of consumed protons in the external medium per one ion of NO sup-inf2 reduced
References
Carlson CA, Ferguson LP, Ingraham JL (1982) Properties of dissimilatory nitrate reductase purified from the denitrifier Pseudomonas aeruginosa. J Bacteriol 151:162–171
Hartree EF (1972) Determination of protein. Modification of the Lowry method that gives a linear photometric response. Anal Biochem 48:422–427
Hochstein LI, Tomlinson GA (1988) The enzymes associated with denitrification. Annu Rev Microbiol 42:231–261
Ingraham JL (1981) Microbiology and genetics of denitrifiers. In: Delwiche CC (ed) Denitrification, nitrification, and atmospheric nitrous oxide. Wiley, New York Chichester Brisbane Toronto, pp 45–65
Kiszkiss DF, Downey RJ (1972) Localization and solubilization of the respiratory nitrate reductase of Bacillus stearothermophilus J Bacteriol 109:803–810
Merck Index (1968) Merck and Co, Inc Rahway, New Jersey, p 744
Meyer TE, Cusanovich MA (1989) Structure, function and distribution of soluble bacterial redox proteins. Biochim Biophys Acta 975:1–28
Nicholas DJD, Nason A (1957) Determination of nitrate and nitrite. In: Colowick SP, Kaplan NO (eds) Methods in enzymology, vol. 3. Academic Press New York, pp 981–984
Scholes P, Mitchell P (1970) Respiration-driven proton translocation in Micrococcus denitrificans. Bioenergetics 1:309–323
Shidara S (1984) Effect of oxygen concentration during culture on denitrifying enzyme system and cytochrome of Bacillus firmus. (In English) J Sci Hiroshima Univ, Ser B Div 2. 19:111–130
Sidransky E, Walter B, Hollocher TC (1978) Studies on the differential inhibition by azide on the nitrite/nitrous oxide level of denitrification. Appl Environ Microbiol 35:247–250
Urata K, Shimada K, Satoh T (1983) Proton translocation associated with denitrification in a photodenitrifier, Rhodopseudomonas sphaeroides forma sp. denitrificans. Plant Cell Physiol 24:501–508
Van't Riet J, Wientjes FB, vanDoorn J, Planta RJ (1979) Purification and characterization of the respiratory nitrate reductase of Bacillus licheniformis. Biochim Biophys Acta 576:347–360
Walter B, Sidransky E, Kristjansson JK, Hollocher TC (1978) Inhibition of denitrification by uncouplers of oxidative phosphorylation. Biochemistry 17:3039–3045
Wientjes FB, Kolk AHJ, Nanninga N, Van't Riet J (1979) Respiratory nitrate reductase: Its localization in the cytoplasmic membrane of Klebsiella aerogenes and Bacillus licheniformis. Eur J Biochem 95:61–67
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Urata, K., Satoh, T. Enzyme localization and orientation of the active site of dissimilatory nitrite reductase from Bacillus firmus . Arch. Microbiol. 156, 24–27 (1991). https://doi.org/10.1007/BF00418182
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00418182